Validation of chronic disease biomarkers in children is of paramount importance especially in the burgeoning arena of pediatric obesity research. Because cardiovascular disease (CVD) is a cumulative process occurring over time, identifying the earliest signs in order to intervene sooner may have a large impact on slowing its progression. The challenge is identifying which obese youth have early vascular problems. Looking to the vascular endothelium for signs of damage is a reasonable approach because of its prominent role in the origins of atherosclerosis. However, accurately quantifying endothelial health in children has proven to be a major challenge. Brachial artery flow-mediated dilation (FMD) is the most commonly-used method to quantify endothelial health in children. However, this technique is not widely applicable because it requires specialized equipment, a highly-trained technician, and results can be highly variable. More direct measures of endothelial cell biology, such as circulating endothelial cells (CEC) and endothelial microparticles (EMP), may offer greater precision in characterizing the state of the endothelium and may identify high-risk individuals. Despite being validated in adults, CEC and EMP have not been formally evaluated in youth. Our primary focus in this study will be the evaluation of CEC and EMP as biomarkers of CVD risk, throughout a spectrum of adiposity in children and adolescents, with a goal of validating CEC and EMP for use in pediatric research studies. We propose a robust study design with a three-tiered approach toward validating these biomarkers: 1) evaluation of the association of CEC and EMP concurrently with CVD risk factors and measures of endothelial function (FMD) and sub-clinical atherosclerosis (carotid intima-media thickness), 2) evaluation of the change in CEC and EMP in response to substantial weight loss in extremely obese adolescents undergoing elective, clinically- indicated bariatric surgery, and 3) longitudinal assessment of the reproducibility of CEC and EMP to inform the design of future pediatric studies using these endpoints. PUBLIC HEALTH RELEVANCE: This project will evaluate the validity of two new markers of vascular damage in children and adolescents with varying levels of body fat in order to assess their potential usefulness in pediatric clinical research studies.